Effects of Macroalgal Chemical Extracts on Spore Behavior of the Antarctic Epiphyte Elachista antarctica Phaeophyceae

Most macroalgal species along the Western Antarctic Peninsula (WAP) are defended against predation, many using chemical defenses. These subtidal communities are also mostly devoid of free living filamentous algae. However, one endo/epiphyte, Elachista antarctica, is found growing exclusively out of the palatable rhodophyte Palmaria decipiens. To understand this unusual and exclusive epiphytization, we tested whether macroalgal secondary metabolites such as those responsible for deterring sympatric grazers, affect the behaviors of the epiphyte's spores. Settlement, germination, and swimming behaviors of the epiphyte's motile spores were quantified in the presence of fractionated lipophilic and hydrophilic extracts of host P. decipiens and other rhodophytes from the shallow subtidal. Host P. decipiens was the only alga tested that did not inhibit spore settlement or germination. We also examined whether extracts from these chemically rich algae affect spore swimming behaviors and found spores to be chemotactically attracted to seawater soluble extract fractions of host P. decipiens. These results indicate that chemosensory behaviors of the epiphyte's spores to metabolites associated with these chemically defended macrophytes can explain this exclusive epiphyte–host interaction.     

Post-translational modification by β-lysylation is required for activity of Escherichia coli elongation factor P (EF-P)

Bacterial elongation factor P (EF-P) is the ortholog of archaeal and eukaryotic initiation factor 5A (eIF5A). EF-P shares sequence homology and crystal structure with eIF5A, but unlike eIF5A, EF-P does not undergo hypusine modification. Recently, two bacterial genes, yjeA and yjeK, encoding truncated homologs of class II lysyl-tRNA synthetase and of lysine-2,3-aminomutase, respectively, have been implicated in the modification of EF-P to convert a specific lysine to a hypothetical β-lysyl-lysine. Here we present biochemical evidence for β-lysyl-lysine modification in Escherichia coli EF-P and for its role in EF-P activity by characterizing native and recombinant EF-P proteins for their modification status and activity in vitro. Mass spectrometric analyses confirmed the lysyl modification at lysine 34 in native and recombinant EF-P proteins. The β-lysyl-lysine isopeptide was identified in the exhaustive Pronase digests of native EF-P and recombinant EF-P isolated from E. coli coexpressing EF-P, YjeA, and YjeK but not in the digests of proteins derived from the vectors encoding EF-P alone or EF-P together with YjeA, indicating that both enzymes, YjeA and YjeK, are required for β-lysylation of EF-P. Endogenous EF-P as well as the recombinant EF-P preparation containing β-lysyl-EF-P stimulated N-formyl-methionyl-puromycin synthesis ～4-fold over the preparations containing unmodified EF-P and/or α-lysyl-EF-P. The mutant lacking the modification site lysine (K34A) was inactive. This is the first report of biochemical evidence for the β-lysylation of EF-P in vivo and the requirement for this modification for the activity of EF-P.     

Oxidative balance and colon and rectal cancer: Interaction of lifestyle factors and genes

Pro-oxidant and anti-oxidant genetic and lifestyle factors can contribute to an individual's level of oxidative stress. We hypothesize that diet, lifestyle and genetic factors work together to influence colon and rectal cancer through an oxidative balance mechanism. We evaluated nine markers for eosinophil peroxidase (EPX), two for myeloperoxidase (MPO), four for hypoxia-inducible factor-1A (HIFIA), and 16 for inducible nitric oxide synthase (NOS2A) in conjunction with dietary antioxidants, aspirin/NSAID use, and cigarette smoking. We used data from population-based case-control studies (colon cancer n=1555 cases, 1956 controls; rectal cancer n=754 cases, 959 controls). Only NOS2A rs2297518 was associated with colon cancer (OR 0.86 95% CI 0.74, 0.99) and EPX rs2302313 and MPO rs2243828 were associated with rectal cancer (OR 0.75 95% CI 0.59, 0.96; OR 0.81 95% CI 0.67, 0.99 respectively) for main effects. However, after adjustment for multiple comparisons we observed the following significant interactions for colon cancer: NOS2A and lutein, EPX and aspirin/NSAID use, and NOS2A (4 SNPs) and cigarette smoking. For rectal cancer we observed the following interactions after adjustment for multiple comparisons: HIF1A and vitamin E, NOS2A (3SNPs) with calcium; MPO with lutein; HIF1A with lycopene; NOS2A with selenium; EPX and NOS2A with aspirin/NSAID use; HIF1A, MPO, and NOS2A (3 SNPs) with cigarette smoking. We observed significant interaction between a composite oxidative balance score and a polygenic model for both colon (p interaction 0.0008) and rectal cancer (p=0.0018). These results suggest the need to comprehensively evaluate interactions to assess the contribution of risk from both environmental and genetic factors.     

The evolution of land plant cilia

Eukaryotic cilia/flagella are ancient organelles with motility and sensory functions. Cilia display significant ultrastructural conservation where present across the eukaryotic phylogeny; however, diversity in ciliary biology exists and the ability to produce cilia has been lost independently on a number of occasions. Land plants provide an excellent system for the investigation of cilia evolution and loss across a broad phylogeny, because early divergent land plant lineages produce cilia, whereas most seed plants do not. This review highlights the differences in cilia form and function across land plants and discusses how recent advances in genomics are providing novel insights into the evolutionary trajectory of ciliary proteins. We propose a renewed effort to adopt ciliated land plants as models to investigate the mechanisms underpinning complex ciliary processes, such as number control, the coordination of basal body placement and the regulation of beat patterns.     

Placenta Stem/Stromal Cell–Derived Extracellular Vesicles for Potential Use in Lung Repair

Many acute and chronic lung injuries are incurable and rank as the fourth leading cause of death globally. While stem cell treatment for lung injuries is a promising approach, there is growing evidence that the therapeutic efficacy of stem cells originates from secreted extracellular vesicles (EVs). Consequently, EVs are emerging as next‐generation therapeutics. While EVs are extensively researched for diagnostic applications, their therapeutic potential to promote tissue repair is not fully elucidated. By housing and delivering tissue‐repairing cargo, EVs refine the cellular microenvironment, modulate inflammation, and ultimately repair injury. Here, the potential use of EVs derived from two placental mesenchymal stem/stromal cell (MSC) lines is presented; a chorionic MSC line (CMSC29) and a decidual MSC cell line (DMSC23) for applications in lung diseases. Functional analyses using in vitro models of injury demonstrate that these EVs have a role in ameliorating injuries caused to lung cells. It is also shown that EVs promote repair of lung epithelial cells. This study is fundamental to advancing the field of EVs and to unlock the full potential of EVs in regenerative medicine.     

Polarization of endosomal SNX27 in migrating and tumor-engaged natural killer cells

Polarization is a critical mechanism for the proper functioning of many cell types. For lymphocytes, it is essential in a variety of processes, including migration from the blood to other tissue sites and vice versa. In NK cells and CTLs, the cytotoxic granule delivery mechanism requires polarization for granule movement to the immunological synapse (IS), in killing tumor and virus-infected cells. Recently, it has become apparent that endosomes are also involved in the cytotoxic mechanism. Using an in vitro conjugation approach, we show that in NK-92 cells, endosomal Sorting Nexin 27 (SNX27) polarizes to the IS during tumor cell engagement in a distinct compartment adjacent to the cytotoxic granules. We also show that SNX27 polarizes to the apical membrane, opposite the uropod, during NK cell migration. These previously unreported results indicate that SNX27 is a participant in NK cell polarization, as a mediator or target of the mechanism.     

Dihydromyricetin Prevents Fetal Alcohol Exposure-Induced Behavioral and Physiological Deficits: The Roles of GABAA Receptors in Adolescence

Fetal alcohol exposure (FAE) can lead to a variety of behavioral and physiological disturbances later in life. Understanding how alcohol (ethanol, EtOH) affects fetal brain development is essential to guide the development of better therapeutics for FAE. One of EtOH’s many pharmacological targets is the γ-aminobutyric acid type A receptor (GABA_AR), which plays a prominent role in early brain development. Acute EtOH potentiates inhibitory currents carried by certain GABA_AR subtypes, whereas chronic EtOH leads to persistent alterations in GABA_AR subunit composition, localization and function. We recently introduced a flavonoid compound, dihydromyricetin (DHM), which selectively antagonizes EtOH’s intoxicating effects in vivo and in vitro at enhancing GABA_AR function as a candidate for alcohol abuse pharmacotherapy. Here, we studied the effect of FAE on physiology, behavior and GABA_AR function of early adolescent rats and tested the utility of DHM as a preventative treatment for FAE-induced disturbances. Gavage administration of EtOH (1.5, 2.5, or 5.0 g/kg) to rat dams on day 5, 8, 10, 12, and 15 of pregnancy dose-dependently reduced female/male offspring ratios (largely through decreased numbers of female offspring) and offspring body weights. FAE (2.5 g/kg) rats tested on postnatal days (P) 25–32 also exhibited increased anxiety and reduced pentylenetetrazol (PTZ)-induced seizure threshold. Patch-clamp recordings from dentate gyrus granule cells (DGCs) in hippocampal slices from FAE (2.5 g/kg) rats at P25-35 revealed reduced sensitivity of GABAergic miniature inhibitory postsynaptic currents (mIPSCs) and tonic current (I_tonic) to potentiation by zolpidem (0.3 μM). Interestingly, potentiation of mIPSCs by gaboxadol increased, while potentiation of I_tonic decreased in DGCs from FAE rats. Co-administration of EtOH (1.5 or 2.5 g/kg) with DHM (1.0 mg/kg) in pregnant dams prevented all of the behavioral, physiological, and pharmacological alterations observed in FAE offspring. DHM administration alone in pregnant rats had no adverse effect on litter size, progeny weight, anxiety level, PTZ seizure threshold, or DGC GABA_AR function. Our results indicate that FAE induces long-lasting alterations in physiology, behavior, and hippocampal GABA_AR function and that these deficits are prevented by DHM co-treatment of EtOH-exposed dams. The absence of adverse side effects and the ability of DHM to prevent FAE consequences suggest that DHM is an attractive candidate for development as a treatment for prevention of fetal alcohol spectrum disorders.     

Loss of microbial diversity in soils is coincident with reductions in some specialized functions

Loss of microbial diversity is considered a major threat because of its importance for ecosystem functions, but there is a lack of conclusive evidence that diversity itself is reduced under anthropogenic stress, and about the consequences of diversity loss. Heavy metals are one of the largest, widespread pollutant types globally, and these represent a significant environmental stressor for terrestrial microbial communities. Using combined metagenomics and functional assays, we show that the compositional and functional response of microbial communities to long‐term heavy metal stress results in a significant loss of diversity. Our results indicate that even at a moderate loss of diversity, some key specialized functions (carried out by specific groups) may be compromised. Together with previous work, our data suggest disproportionate impact of contamination on microbes that carry out specialized, but essential, ecosystem functions. Based on these findings, we propose a conceptual framework to explicitly consider diversity of functions and microbial functional groups to test the relationship between biodiversity and soil functions.     

Molecular Dynamics Study of the Opening Mechanism for DNA Polymerase I

During DNA replication, DNA polymerases follow an induced fit mechanism in order to rapidly distinguish between correct and incorrect dNTP substrates. The dynamics of this process are crucial to the overall effectiveness of catalysis. Although X-ray crystal structures of DNA polymerase I with substrate dNTPs have revealed key structural states along the catalytic pathway, solution fluorescence studies indicate that those key states are populated in the absence of substrate. Herein, we report the first atomistic simulations showing the conformational changes between the closed, open, and ajar conformations of DNA polymerase I in the binary (enzyme∶DNA) state to better understand its dynamics. We have applied long time-scale, unbiased molecular dynamics to investigate the opening process of the fingers domain in the absence of substrate for B. stearothermophilis DNA polymerase in silico. These simulations are biologically and/or physiologically relevant as they shed light on the transitions between states in this important enzyme. All closed and ajar simulations successfully transitioned into the fully open conformation, which is known to be the dominant binary enzyme-DNA conformation from solution and crystallographic studies. Furthermore, we have detailed the key stages in the opening process starting from the open and ajar crystal structures, including the observation of a previously unknown key intermediate structure. Four backbone dihedrals were identified as important during the opening process, and their movements provide insight into the recognition of dNTP substrate molecules by the polymerase binary state. In addition to revealing the opening mechanism, this study also demonstrates our ability to study biological events of DNA polymerase using current computational methods without biasing the dynamics.     

EGb761 Provides a Protective Effect against Aβ1-42 Oligomer-Induced Cell Damage and Blood-Brain Barrier Disruption in an In Vitro bEnd.3 Endothelial Model

Alzheimer’s disease (AD) is the most common form of senile dementia which is characterized by abnormal amyloid beta (Aβ) accumulation and deposition in brain parenchyma and cerebral capillaries, and leads to blood-brain barrier (BBB) disruption. Despite great progress in understanding the etiology of AD, the underlying pathogenic mechanism of BBB damage is still unclear, and no effective treatment has been devised. The standard Ginkgo biloba extract EGb761 has been widely used as a potential cognitive enhancer for the treatment of AD. However, the cellular mechanism underlying the effect remain to be clarified. In this study, we employed an immortalized endothelial cell line (bEnd.3) and incubation of Aβ_1–42 oligomer, to mimic a monolayer BBB model under conditions found in the AD brain. We investigated the effect of EGb761 on BBB and found that Aβ_1–42 oligomer-induced cell injury, apoptosis, and generation of intracellular reactive oxygen species (ROS), were attenuated by treatment with EGb761. Moreover, treatment of the cells with EGb761 decreased BBB permeability and increased tight junction scaffold protein levels including ZO-1, Claudin-5 and Occludin. We also found that the Aβ_1–42 oligomer-induced upregulation of the receptor for advanced glycation end-products (RAGE), which mediates Aβ cytotoxicity and plays an essential role in AD progression, was significantly decreased by treatment with EGb761. To our knowledge, we provide the first direct in vitro evidence of an effect of EGb761 on the brain endothelium exposed to Aβ_1–42 oligomer, and on the expression of tight junction (TJ) scaffold proteins and RAGE. Our results provide a new insight into a possible mechanism of action of EGb761. This study provides a rational basis for the therapeutic application of EGb761 in the treatment of AD.